Abstract
Protein structure determination by X-ray crystallography is dependent on obtaining a single protein crystal suitable for diffraction data collection. Due to this requirement, protein crystallization represents a key step in protein structure determination. The conditions for protein crystallization have to be determined empirically for each protein, making this step also a bottleneck in the structure determination process. Typical protein crystallization practice involves parallel setup and monitoring of a considerable number of individual protein crystallization experiments (also called crystallization trials). In these trials the aliquots of purified protein are mixed with a range of solutions composed of a precipitating agent, buffer, and sometimes an additive that have been previously successful in prompting protein crystallization. The individual chemical conditions in which a particular protein shows signs of crystallization are used as a starting point for further crystallization experiments. The goal is optimizing the formation of individual protein crystals of sufficient size and quality to make them suitable for diffraction data collection. Thus the composition of the primary crystallization screen is critical for successful crystallization.
Systematic analysis of crystallization experiments carried out on several hundred proteins as part of large-scale structural genomics efforts allowed the optimization of the protein crystallization protocol and identification of a minimal set of 96 crystallization solutions (the “TRAP” screen) that, in our experience, led to crystallization of the maximum number of proteins.
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References
Chayen NE (2004) Turning protein crystallization from an art into a science. Curr Opin Struct Biol 14(5):577–583
McPherson A (2004) Introduction to protein crystallization. Methods 34(3):254–265
Chayen NE, Saridakis E (2008) Protein crystallization: from purified protein to diffraction-quality crystal. Nat Methods 5(2):147–153
Bergfors T (2009) Protein crystallization. In: Tsigelny IF (ed) IUL biotechnology, 2nd edn. International University Line, La Jolla
Ochi T et al (2009) Perspectives on protein crystallization. Prog Biophys Mol Biol 101(1–3):56–63
Bolanos-Garcia VM, Chayen NE (2009) New directions in conventional methods of protein crystallization. Prog Biophys Mol Biol 101(1–3):3–12
McPherson A (1990) Current approaches to macromolecular crystallization. Eur J Biochem 189(1):1–23
Saridakis E, Chayen NE (2003) Systematic improvement of protein crystals by determining the supersolubility curves of phase diagrams. Biophys J 84(2 Pt 1):1218–1222
McPherson A (1985) Crystallization of proteins by variation of pH or temperature. Methods Enzymol 114:125–127
Enrico A, Stura GRN, Wilson IA (1992) Strategies in the crystallization of glycoproteins and protein complexes. J Cryst Growth 122(1–4):273–285
Blow DM et al (1994) Control of nucleation of protein crystals. Protein Sci 3(10):1638–1643
Benvenuti M, Mangani S (2007) Crystallization of soluble proteins in vapor diffusion for X-ray crystallography. Nat Protoc 2(7):1633–1651
Jancarik J, Kim S-H (1991) Sparse matrix sampling: a screening method for crystallization of proteins. J Appl Cryst 24:409–411
Bulek AM et al (2012) TCR/pMHC optimized protein crystallization screen. J Immunol Methods 382(1–2):203–210
Grimm C et al (2010) A crystallization screen based on alternative polymeric precipitants. Acta Crystallogr D Biol Crystallogr 66(Pt 6):685–697
Berry IM et al (2006) SPINE high-throughput crystallization, crystal imaging and recognition techniques: current state, performance analysis, new technologies and future aspects. Acta Crystallogr D Biol Crystallogr 62(Pt 10):1137–1149
Chayen NE, Saridakis E (2002) Protein crystallization for genomics: towards high-throughput optimization techniques. Acta Crystallogr D Biol Crystallogr 58(Pt 6 Pt 2):921–927
Sauder MJ et al (2008) High throughput protein production and crystallization at NYSGXRC. Methods Mol Biol 426:561–575
Stewart L, Clark R, Behnke C (2002) High-throughput crystallization and structure determination in drug discovery. Drug Discov Today 7(3):187–196
Sugahara M et al (2008) High-throughput crystallization-to-structure pipeline at RIKEN SPring-8 center. J Struct Funct Genomics 9(1–4):21–28
Page R, Stevens RC (2004) Crystallization data mining in structural genomics: using positive and negative results to optimize protein crystallization screens. Methods 34(3):373–389
Hui R, Edwards A (2003) High-throughput protein crystallization. J Struct Biol 142(1):154–161
Kimber MS et al (2003) Data mining crystallization databases: knowledge-based approaches to optimize protein crystal screens. Proteins 51(4):562–568
Acknowledgements
This work was supported with federal funds from the National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services, under Contract Nos. HHSN272200700058C and HHSN272201200026C.
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Skarina, T., Xu, X., Evdokimova, E., Savchenko, A. (2014). High-Throughput Crystallization Screening. In: Anderson, W.F. (eds) Structural Genomics and Drug Discovery. Methods in Molecular Biology, vol 1140. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-0354-2_12
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DOI: https://doi.org/10.1007/978-1-4939-0354-2_12
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